The goal of the proposed research is to understand the processes involved in the inhibition of cell differentiation produced by the expression of the transforming gene of Rous Sarcoma Virus (RSV). The method of approach will be to analyze several biochemical and cellular parameters of a rat myogenic cell line (L6E9) which has been transformed with an RSV with a temperature sensitive (ts) mutation on the "src" gene. First we will determine the phenotype of the transformed cells at permissive and non-permissive temperatures. We will study the changes produced by src on the expression of the major contractile protein, myosin heavy chain (MHC). The effects of src on MHC will be compared with its effects on a "house-keeping" function. We will study the expression of these two host genes at the protein and mRNA level to determine whether the changes in phenotype produced by cell transformation occurs at the transcriptional, post-transcriptional, translational, or post-translational level. MHC and house-keeping mRNA sequences will be assayed by DNA-RNA hybridization methods using cDNA sequences cloned in bacterial plasmids. Studies with whole cells will be complemented with experiments designed to measure transcription in isolated nuclei. Second, we will initiate experiments to determine what cellular parameters are involved in the production of the molecular phenotype. To this end we will determine whether the inhibitory effect of src on MHC expression is through a change on the growth properties of the myoblasts or by directly affecting the differentiation program.